PROGESTERONE SECRETION AND PROLIFERATION IN CULTURED RABBIT GRANULOSA-CELLS UNDER CONDITIONS OF BETA-D-XYLOSIDE-INDUCED INHIBITION OF PROTEOGLYCAN SYNTHESIS

Proteoglycans present in follicular fluid are synthesized by granulosa cells under gonadotropin control. An inhibitor of proteoglycan synthe sis, p-nitrophenyl-beta-D-xyloside (beta-D-xyloside) was used as a pro be to study rabbit granulosa cell steroidogenesis and proliferation un der abrogated proteoglycan synthesis. Granulosa cells isolated from ra bbit preovulatory follicles were cultured 24 h in Minimum Essential Me dium plus 2.5% fetal calf serum in the presence of absence of beta-D-x yloside and were then treated with FSH or dibutyryl cAMP (db-cAMP) alo ne or in combination with beta-D-xyloside for a further 24 h. The expo sure for 48 h of granulosa cells to 1 mM beta-D-xyloside in the absenc e or presence of FSH inhibited proteoglycan synthesis and increased th e amount of glycosaminoglycans (GAG). FSH-stimulated progesterone prod uction was significantly correlated only with proteoglycan synthesis a nd not with GAG production. The addition of various concentrations of beta-D-xyloside (0.1-4 mM) for 48 h to granulosa cells induced a dose- dependent inhibition of FSH-stimulated progesterone secretion and [H-3 ]thymidine incorporation into DNA. beta-D-Xyloside concentrations lowe r than 1 mM induced an inhibition of FSH-stimulated progesterone secre tion but had no significant effect on FSH-induced proliferation. One m illimolar beta-D-xyloside did not modify basal progesterone production , but in the presence of various doses (0.1-2.5 ng/ml) of FSH of hCG ( 0.1-1 IU/ml) it exerted a significant inhibitory effect on steroid sec retion. Fifty percent inhibition was obtained for doses of FSH above 0 .5 ng/ml. When cells were stimulated by db-cAMP instead of FSH, the in hibition of progesterone synthesis induced by 1 mM beta-D-xyloside was slightly significant. This dose of beta-D-xyloside markedly inhibited FSH-induced cAMP production. Thus, it seems that the major consequenc e of proteoglycan synthesis abrogation was at a step beyond cAMP synth esis. These results suggest a role of proteoglycan metabolism in maint aining important functions of granulosa cell steroidogenesis and proli feration.